The long-term objective of this proposal is to provide state-of-the-art confocal imaging capabilities to advance NIH-funded projects in the Cardio regenerative Research Program and Cellular and Molecular Arrhythmia Research Program at the University of Wisconsin-Madison. Ongoing research projects by investigators in these groups currently utilize confocal microscopy extensively, but the available dedicated instrument (Bio-Rad Micro radiance) has recently been retired (10 years old) because of lack of service support and high costs associated with additional repair. Furthermore, technological advances and new features in contemporary confocal microscopes provide a significant advance in imaging capabilities. A fundamental need for research in cardio regenerative medicine is to be able to track transplanted cells and determine their fate in cardiac muscle. The five channel detection provided by the FV1000 will assist cardio regenerative research projects by allowing the simultaneous detection of multiple distinct cell lineage-specific proteins to more clearly demarcate distinct cell types in repaired and regenerated myocardium. Furthermore, the spectral detection system provided by the Olympus FV1000 will allow a more accurate detection of specific fluorophores, minimizing cross-talk. Critical questions in basic arrhythmia research are increasingly focused on determining the trafficking of ion channel proteins and associated proteins to the surface membrane and determining the precise sub compartmentalization of these molecules in cardiomyocytes. The proposed Olympus FV1000 will enable dynamic tracking of the membrane trafficking of ion channels and associated proteins by enabling fluorescence recovery after photo bleaching (FRAP) experiments using the SIM (simultaneous laser light stimulation and imaging) functionality of the system. Additionally, the ability to perform combined cellular electrophysiology and confocal imaging experiments greatly enhances the functional information that can be obtained by these experiments using techniques such as dynamic fluorescence resonance energy transfer (FRET) during voltage clamp experiments and calcium imaging with simultaneous voltage clamp studies. Comparisons with other instruments on the market suggested that the Olympus FV1000 has the proven functionality at a competitive price for the specified needs with excellent service available. The necessary space, administrative support, supervision, and technical expertise are already in place. An advisory board consisting not only of users but also of experts in confocal imaging and cardiac biology will provide oversight. Institutional support from the School of Medicine and Public Health, Department of Medicine, and Division of Cardiovascular Medicine will provide a microscope manager and funds needed for maintenance and repair. Overall, the requested Olympus FV1000 microscope will significantly advance multiple research projects improving current imaging data acquisition as well as providing new functionality that will enable powerful new applications not possible on available equipment. PUBLIC HEALTH REVELANCE: Heart disease remains the most common cause of death in the United States. The requested confocal microscope will provide a powerful tool for research projects aimed at better understanding the cellular and molecular mechanisms of arrhythmias, a major cause of illness and sudden death. In addition, this microscope will enable pioneering research focused on identifying new strategies to repair and regenerate cardiac muscle following injury such as myocardial infarction.